在NifEN上优化FeMoco成熟的优化
Janice M Yoshizawa1, Michael A Blank, Aaron W Fay
1Department of Molecular Biology and Biochemistry, University of California, Irvine, California 92697, USA.
Journal of the American Chemical Society
|June 12, 2009
概括
优化的二尼特度 (20毫米) 增强了与NifEN支架蛋白结合的铁合因子 (FeMoco) 前体的成熟. 这种改进的成熟允许NifEN完全激活FeMoco缺少的MoFe蛋白,模仿孤立的FeMoco活性.
科学领域:
- 生物化学 生物化学
- 生物有机化学 生物有机化学
- 酶学 是一种酶学.
背景情况:
- 酶酶,特别是Mo-酶,对于将大气中的二转化为氨非常重要.
- 铁辅因子 (FeMoco) 是这种降解的活性位点.
- 尼芬作为支架蛋白,对于FeMoco的正确生物合成和成熟至关重要.
研究的目的:
- 为了优化与NifEN结合的FeMoco前体的体外成熟.
- 调查二尼酸盐度对FeMoco成熟和NifEN激活MoFe蛋白的能力的影响.
- 为了描述优化NiFEN结合的FeMoco的结构和特性.
主要方法:
- 在体外生化试验测量MoFe蛋白激活.
- 电子偏磁共振 (EPR) 谱学分析FeMoco活性部位.
- 进行X射线吸收光谱 (XAS) 和扩展X射线吸收细结构 (EXAFS) 分析以进行结构性特征.
主要成果:
- 使用20mM迪西奥尼特进行了优化成熟,结果在NiFEN结合的FeMoco中形成了一个紧密组织的Mo位点.
- 经过最佳成熟后,NifEN结合的FeMoco激活了FeMoco缺少的MoFe蛋白,其程度与分离的FeMoco相美.
- EPR和XAS/EXAFS数据显示,NifEN中的FeMoco构造类似于MoFe蛋白中的FeMoco构造.
结论:
- FeMoco成熟对迪尼特度高度敏感,突出了氧化还原化学的关键作用.
- 优化的二化物条件为了解FeMoco生物合成中的体内电子转移机制提供了一个模型.
- 这项研究为在体外使用NifEN支架生成完全功能化的FeMoco提供了一条途径.
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